(1) Field of the Invention
The present invention generally relates to inhibitors of protein action. More specifically, the invention is directed to the identification of small molecule inhibitors of BCL6.
(2) Description of the Related Art
The BTB domain (also known as the POZ domain) is a common protein-protein interaction motif that is found in over 180 human proteins (Stogios et al., 2005). The domain has been found in proteins that are implicated in many biological processes, including central nervous system development, oocyte maturation, eye development, hematopoiesis, apoptosis, immunity, and protein degradation. BTB domain proteins are widely represented in eukaryotic genomes, and are roughly as abundant as SH3 domain proteins. One major class of BTB proteins consists of a single N-terminal BTB domain, a middle linker region, and a set of C-terminal C2H2 zinc-finger domains, and at least 43 such BTB-ZF genes have been identified in the human genome (Srogios et al., 2005). These proteins are also known as the POK proteins (Maeda et al., 2005). Many BTB-ZF proteins have been implicated in cancer, including BCL6 (Albagli-Curiel, 2003; Polo et al., 2004; Cattoretti et al., 2005), PLZF (Costoya and Pandolfi, 2001; McConnell et al., 2003), LRF/Pokemon (Maeda et al., 2005), HIC1 (Chen et al., 2004; Pinte et al., 2004), Miz-1 (Peulert et al., 1997; Phan et al., 2005), and Kaiso (Prokhortchouk et al., 2006). In general, the zinc fingers from these proteins are responsible for binding to specific regulatory sites on the DNA, while the BTB domain is a protein-protein interaction module that dimerizes and functions to modulate the transcriptional activity of the factors.
Both the BCL6 and PLZF proteins consist of an N-terminal BTB domain, followed by a central region of several hundred residues that are predicted to have little or no fixed 3D structure, and end with a series of C2H2-type zinc finger domains at the C-terminus. This general type of architecture is seen in 43 of the over 200 known human BIB domain proteins (GGP and P. J. Stogios, xtal.uhnres.utoronto.ca/prive/btb.html). A second major class of BTB domain proteins contain C-terminal ketch β-propeller repeats, and many of these are thought to be involved in cytoskeletal functions, although some of these are involved in transcription regulation (Adams et al., 2000). The core BTB domain fold is also found in the T1 domain of voltage-gated channels (Kreusch et al., 1998), and in the ElonginC/Skp1 proteins (Stebbins et al., 1999).
Despite the architectural similarity of the BTB/zinc finger transcription factors, these can function as repressors, activators, or both and the BIB domain plays a central role in these activities (Kaplan and Calame, 1997; Kobayashi et al., 2000; Mahmoudi et al., 2002). The majority of BTB/zinc finger proteins, however, are thought to be transcriptional repressors, and several of these mediate their effects through the recruitment of histone deacetylase complexes. Thus, in BCL6, the BIB domain mediates interactions with the SMRT, N-CoR, BCoR and mSin3A corepressors, as well as with histone deacetylase 1 (HDAC-1), and repression is relieved with HDAC inhibitors (David et al., 1998; Dhordain et al., 1997; Dhordain et al., 1998; Grignani et al., 1998; Guidez et al., 1998; He et al., 1998; Hong et al., 1997; Huynh and Bardwell, 1998; Huynh et al., 2000; Lin et al., 1998; Wong and Privalsky, 1998). The recruitment of a histone deacetylase complex is not a universal property of the BTB domain, as evidenced by the fact that the BTB domains of HIC1 and gFBP-B do not interact with these factors (Deltour et al., 1999). Thus, it is clear that distinct mechanisms are used by different BTB domains in order to carry out a variety of biological effects.
In the B-cell lineage, the BCL6 protein is expressed in germinal center (GC) B-cells, but not in pre-B cells or in differentiated progenies such as plasma cells. Because BCL6 expression is tightly regulated during lymphoid differentiation, its down-regulation in post-GC B-cells may be necessary for further plasma/memory cell differentiation. Some of the more notable genes that are repressed by BCL6 include the B lymphocyte-induced maturation protein (blimp-1), a transcriptional repressor of c-myc which plays a key role in differentiation of B-cells to plasma cells (Shaffer et al., 2002), the cell cycle control genes p27kip1 and cyclin D2 (Shaffer et al., 2000), the programmed cell death-2 protein (PDCD2) (Baron et al., 2002), and B7-1/CD80 (Niu et al., 2003). Chromosomal translocations upstream of the BCL6 gene are observed in approximately 30-40% of diffuse large B-cell lymphomas (DLBCL) and in 5-14% of follicular lymphomas (FL) (Kuppers and Dalla-Favera, 2001; Niu, 2002; Ye, 2000). In addition, the promoter region of BCL6 is targeted by somatic hypennutation in GC B-cells (Pasqualucci et al., 2003; Shen et al., 1998; Wang et al., 2002). Thus, a B-cell with an activated BCL6 gene may be trapped at the GC stage due to the repression of differentiation and cell-cycle control proteins (Calame et al., 2003; Dent et al., 2002; Fearon et al., 2001; Staudt, 2002). In addition to its role in lymphoid cells, BCL6 represses the expression of the chemokines MCP-1, MCP3 and MRP-1 in macrophages and is an important negative regulator of TH-2 type inflammation (Toney et al., 2000).
The BCL6 site of corepressor binding has been identified, and peptides having the sequence of the corepressor binding site inhibit corepressor binding to BCL6. That inhibition causes apoptosis of B-cell lymphoma cells expressing BCL6. See WO 2005/058939 A2. In that work, a recombinant BCL6 peptide inhibitor (BPI) was designed based on the BCL6/SMRT crystal. BPI could disrupt the formation of the BCL6/SMRT complex on target gene promoters, reactivate BCL6 target gene expression in lymphoma cells and could phenocopy the BCL6 null phenotype in vivo (i.e. loss of germinal center formation in response to T-cell dependent antigens) (Polo et al., 2004). As alluded to above, BPI has potent anti-lymphoma activity, and killed BCL6 positive DLBCL and BL cell lines in vitro and in vivo (in xenotransplants) but had no effect on BCL6 negative lymphoma cell lines (Polo et al., 2004). Moreover, there was no toxicity observed in animals injected daily even with up to 1.5 mg of BPI per day (Polo et al., 2004; unpublished data). Those results indicate that BCL6 is an excellent therapeutic target and that lateral groove blockade is a potent and specific strategy for abrogating its activity in lymphoma cells.
Based on the above, it is apparent that small molecule inhibitors of BCL6 would be useful for cancer treatments. The present invention describes such BCL6 inhibitors.